Novel reporter constructs for compound screening

Abstract

The instant description provides reporter constructs, transgenic cells, and transgenic organisms and methods for identifying agents that can regulate gene expression and improve plant performance and yield. Compounds that increase plant performance or yield are identified by contacting a test compound with a plant cell that comprises a target promoter sequence operably linked to a polynucleotide sequence encoding a DNA sequence-specific transactivator, and a reporter polynucleotide that is operably linked to a promoter sequence that is recognized by the DNA sequence-specific transactivator. The target promoter sequence can be recognized by a transcriptional regulatory polypeptide capable of modulating specific signaling pathways that enhance plant performance or yield.

What is claimed is: 
     
         1 . A method of screening for useful compounds comprising the steps of:
 (a) contacting at least one test compound with a transgenic cell comprising a target promoter sequence that is operably linked to a polynucleotide sequence that encodes a DNA sequence-specific transactivator, and a reporter polynucleotide that is operably linked to a promoter sequence that can be recognized by the DNA sequence-specific transactivator; and   (b) selecting a compound that alters the reporter gene activity relative to controls.   
     
     
         2 . The method of  claim 1 , wherein the DNA sequence-specific transactivator is a translational fusion of a DNA binding domain (DBD) and a transcriptional activation domain (AD);
 wherein the AD comprises at least one sequence selected from the group consisting of SEQ ID NOs: 20, and 22-26; and   wherein the DBD comprises at least one sequence selected form the group consisting of SEQ ID NOs: 16 and 18.   
     
     
         3 . The method of  claim 1 , wherein the reporter polynucleotide encodes a polypeptide that is selected from the group of green fluorescent protein, luciferase, chloramphenicol transferase, and glucuronidase. 
     
     
         4 . The method of  claim 1 , wherein the target promoter sequence is selected from the group consisting of an AT5G52310 (RD29A) promoter, an At5g52300 promoter, an AT1G16850 promoter, an At3g46230 promoter, an AT1G52690 promoter, an At2g37870 promoter, an AT5G43840 promoter, an At5g66780 promoter, an At3g17520 promoter, an At4g09600 promoter, an AT1G15125 promoter, an AT1G13300 promoter, an AT2G48080 promoter, an AT3G25790 promoter, an AT5G10210 promoter, and an AT5G19970 promoter. 
     
     
         5 . The method of  claim 1 , wherein the transgenic cell further comprises a polynucleotide sequence that encodes an additional DNA sequence-specific transactivator,
 wherein transcription of the polynucleotide sequence encoding the additional DNA-sequence-specific transactivator is under the control of a promoter sequence that is recognized by the first DNA-sequence-specific transactivator; and wherein the additional DNA-sequence-specific transactivator regulates transcription of the reporter gene through binding of its cognate sequence that is operably linked to the reporter polynucleotide.   
     
     
         6 . The method of  claim 1  further comprising the step of:
 (c) contacting a plant with the selected compound and detecting a modified trait in the plant relative to controls. 
 
     
     
         7 . The method of  claim 6 , wherein the modified trait is selected from the group of increased desiccation or drought tolerance, increased nitrogen use efficiency, and increased disease resistance.